Stephen W. White

Crystal structure of prokaryotic ribosomal protein L9: a bi-lobed RNA-binding protein.

The crystal structure of protein L9 from the Bacillus stearothermophilus ribosome has been determined at 2.8 A resolution using X‐ray diffraction methods. This primary RNA‐binding protein has a highly elongated and unusual structure consisting of two separated domains joined by a long exposed alpha‐helix. Conserved, positively charged and aromatic amino acids on the surfaces of both domains probably represent the sites of specific interactions with 23S rRNA. Comparisons with other prokaryotic L9 sequences show that while the length of the connecting alpha‐helix is invariant, the sequence within the exposed central region is not conserved. This suggests that the alpha‐helix has an architectural role and serves to fix the relative separation and orientation of the N‐ and C‐terminal domains within the ribosome. The N‐terminal domain has structural homology to the smaller ribosomal proteins L7/L12 and L30, and the eukaryotic RNA recognition motif (RRM).

Structural evidence for specific S8–RNA and S8–protein interactions within the 30S ribosomal subunit: ribosomal protein S8 from Bacillus stearothermophilus at 1.9 å resolution

BACKGROUND Prokaryotic ribosomal protein S8 is an important RNA-binding protein that occupies a central position within the small ribosomal subunit. It interacts extensively with 16S rRNA and is crucial for the correct folding of the central domain of the rRNA. S8 also controls the synthesis of several ribosomal proteins by binding to mRNA. It binds specifically to very similar sites in the two RNA molecules. RESULTS S8 is divided into two tightly associated domains and contains three regions that are proposed to interact with other ribosomal components: two potential RNA-binding sites, and a hydrophobic patch that may interact with a complementary hydrophobic region of S5. The N-terminal domain fold is found in several proteins including two that bind double-stranded DNA. CONCLUSIONS These multiple RNA-binding sites are consistent with the role of S8 in organizing the central domain and agree with the latest models of the 16S RNA which show that the S8 location coincides with a region of complicated nucleic-acid structure. The presence in a wide variety of proteins of a region homologous to the N-terminal domain supports the idea that ribosomal proteins must represent some of the earliest protein molecules.

The crystal structure of ribosomal protein L14 reveals an important organizational component of the translational apparatus

BACKGROUND Detailed structural information on ribosomal proteins has increased our understanding of the structure, function and evolution of the ribosome. L14 is one of the most conserved ribosomal proteins and appears to have a central role in the ribonucleoprotein complex. Studies have indicated that L14 occupies a central location between the peptidyl transferase and GTPase regions of the large ribosomal subunit. RESULTS The crystal structure of L14 from Bacillus stearothermophilus has been solved using a combination of isomorphous replacement and multiwavelength anomalous dispersion (MAD) methods. The structure comprises a five-stranded beta-barrel, a C-terminal loop region that contains two small alpha-helices, and a beta-ribbon that projects from the beta-barrel. An analysis of the structure and the conserved amino acids reveals three surface patches that probably mediate L14-RNA and L14-protein interactions within the ribosome. CONCLUSIONS The accepted role of ribosomal proteins is to promote the folding and stabilization of ribosomal RNA. The L14 structure is consistent with this notion, and it suggests that the RNA binds in two sites. One RNA-binding site appears to recognize a distinct region of ribosomal RNA during particle assembly. The second site is smaller and may become occupied during the later compaction of the RNA. The surface hydrophobic patch is a likely site of protein-protein interaction, possibly with L19.

Helicase–Contrahelicase Interaction and the Mechanism of Termination of DNA Replication

Termination of DNA replication at a sequence-specific replication terminus is potentiated by the binding of the replication terminator protein (RTP) to the terminus sequence, causing polar arrest of the replicative helicase (contrahelicase activity). Two alternative models have been proposed to explain the mechanism of replication fork arrest. In the first model, the RTP-terminus DNA interaction simply imposes a polar barrier to helicase movement without involving any specific interaction between the helicase and the terminator proteins. The second model proposes that there is a specific interaction between the two proteins, and that the DNA-protein interaction both restricts the fork arrest to the replication terminus and determines the polarity of the process. The evidence presented in this paper strongly supports the second model.

Crystallization of the globular domain of histone H5.

The globular domain of histone H1/H5 binds to the nucleosome and is crucial for the formation of chromatin higher order structure. We have expressed in Escherichia coli a gene that codes for the globular domain of H5. The protein produced in E. coli is functional in nucleosome binding assays. We have obtained crystals of the protein that diffract to beyond 2.5 A (1 A = 0.1 nm) resolution. The crystals are orthorhombic with unit cell dimensions of a = 80.1 A, b = 67.5 A and c = 38.0 A.

Proteins of the Bacillus stearothermophilus ribosome: A 5 Å structure analysis of protein S5

The structure of protein S5 from the small subunit of the Bacillus stearothermophilus ribosome is described to a resolution of 5 Å. The molecular boundary is visible and shows the protein to be essentially compact although slightly elongated in one dimension.

Proteins of the Bacillus stearothermophilus ribosome: Crystallization of protein L6

Crystals of ribosomal protein L6 from Bacillus stearothermophilus suitable for high resolution structural studies have been obtained. Crystals are hexagonal with space group P6122 (or the enantiomorph P6522) and cell dimensions a = b = 72.7 Å, c = 124.9 Å. A search for heavy atom derivatives is in progress.

Proteins of the Bacillus stearothermophilus ribosome: The structure of L6 at 6 Å resolution

The structure of protein L6 from the large subunit of the Bacillus stearothermophilus ribosome is described at a nominal resolution of 6 Å. The protein appears to be compact, with an axial ratio significantly below 2:1.

Proteins of the Bacillus stearothermophilus ribosome. A low resolution crystal analysis of protein L30.

The 5 Å resolution crystal structure analysis of ribosomal protein L30 from Bacillus stearothermophilus is described. The molecule is shown to be compact and extend to about 25–30 Å in each dimension.

The structure of DNA binding protein II at 6 Å resolution

The structure of DNA binding protein II from Bacillus stearothermophilus is described at 6 Å resolution. The molecule exists as the dimer in our crystals, and there are 3 independent dimers related by non‐crystallographic symmetry in the unit cell. The dimer is compact and globular with dimensions of about 32 × 35 × 39 Å3.